Tang et al.

BMC Musculoskeletal Disorders (2018) 19:445

https://doi.org/10.1186/s12891-018-2373-3

RESEARCH ARTICLE Open Access

Latent infection of low-virulence anaerobic

bacteria in degenerated lumbar
intervertebral discs
Guoqing Tang, Zhuo Wang, Ji Chen, Zhengshi Zhang, Hongbin Qian and Yong Chen*

Abstract
Background: The existence of latent low-virulence anaerobic bacteria in degenerated intervertebral discs (IVDs)
remains controversial. In this study, the prevalence of low-virulence anaerobic bacteria in degenerated IVDs was
examined, and the correlation between bacterial infection and clinical symptoms was analysed.
Methods: Eighty patients with disc herniation who underwent discectomy were included in this study. Under a
stringent protocol to ensure sterile conditions, 80 disc samples were intraoperatively retrieved and subjected to
microbiological culture. Meanwhile, tissue samples from the surrounding muscle and ligaments were harvested and
cultured as contamination markers. The severity of IVD degeneration and the prevalence of Modic changes (MCs)
were assessed according to preoperative MRI analysis.
Results: Of the 80 cultured discs, 54 were sterile, and 26 showed the presence of bacteria: Propionibacterium acnes
(21 cases) and coagulase-negative staphylococci (5 cases). MRI revealed that the presence of bacteria was significantly
associated with MCs (P<0.001). However, there was no significant association between bacterial infection and the
severity of IVD degeneration (P = 0.162).
Conclusions: Our findings further validated the presence of low-virulence anaerobic bacteria in degenerated IVDs, and
P. acnes was the most frequent bacterium. In addition, the latent infection of bacteria in IVDs was associated with
Modic changes. Therefore, low-virulence anaerobic bacteria may play a crucial role in the pathophysiology of MCs and
lumbar disc herniation.
Keywords: Low-virulence anaerobic bacteria, Modic changes, Propionibacterium acnes, Coagulase-negative
staphylococci, Disc herniation

Background related to disc inflammation and mechanical injury. Over

Disc degeneration is a multifaceted chronic process that
can lead to herniation, radiculopathy, myelopathy, spinal
stenosis, and degenerative spondylolisthesis and that alters
intervertebral disc structure and function, potentially result-
ing in acute or chronic pain [1]. However, there is a poor
correlation between degenerate disc imaging findings and
back pain. While radicular symptoms are often related to
nerve root compression by a prolapsed disc, it is not un-
common to observe a discrepancy between the degree of
disc prolapse and pain severity [2]. This incongruity be-
tween clinical symptoms and imaging findings is likely
* Correspondence: chenyongspine@163.com
Orthopedic Center, Kunshan Hospital of Traditional Chinese Medicine,
Kunshan, Suzhou 215000, China
the last decade, emerging evidence suggests that low-grade
infection is another possible factor [3, 4].
Stirling et al. performed microbiological culturing of
intervertebral disc material from 36 patients who under-
went micro-discectomy for severe sciatica and found
that 53% were positive for Propionibacterium acnes [3].
Furthermore, serological testing of 140 patients with sci-
atica revealed that 31% had increased titres of serum im-
munoglobulin G to the lipid S antigen. Their findings
suggested that microbial infection could be responsible
for sciatica in disc herniation. Additionally, a recent
RCT demonstrated that antibiotic treatment was effect-
ive for the treatment of chronic low back pain, implying
that disc herniation could be secondary to infection [5].

© The Author(s). 2018 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0
International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and
reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to
the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver
(http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
Tang et al. BMC Musculoskeletal Disorders (2018) 19:445
Approximately 70% of studies report that nonspecific
low back pain is positively associated with Modic
changes (MCs), i.e., bone oedema in vertebrae, with odds
ratios ranging from 2.0 to 19.9 [4, 6]. Moreover, Albert
et al. observed new MC development within 1–2 years
after discectomy in 80% of patients with anaerobic bac-
teria detected in their discs, compared to only 44% of
patients with negative cultures [4]. Urquhart et al. per-
formed a systematic review of 11 published studies on
this topic and found moderate evidence indicating that
low-virulence bacteria play a role in disc herniation with
low back pain and that bacterial presence is related to
MCs associated with disc herniation [7].
Nevertheless, there are still many challenges regarding
the topic of low anaerobic bacterial infection in interver-
tebral discs. For example, there is still not enough clin-
ical research throughout the world to replicate the
results and verify the conclusion. Additionally, the lack
of reliable contamination control during disc harvest or
culture in some studies renders the results less accurate.
Additionally, the correlation among bacterial infection,
Modic changes and low back pain needs to be investi-
gated with better clinical data. Therefore, the aim of the
present study was to analyse the possible presence of
bacteria in intervertebral disc biopsies of patients who
underwent surgery for lumbar disc herniation and to
examine the association between bacterial presence and
MCs. Verification of such a link could result in a con-
ceptual paradigm shift and usher in a new era of lumbar
disc problem management.
Methods
Patients
Eighty consecutive patients from January 2017 to August
2017 who underwent discectomy due to a single level of
lumbar disc herniation were included in this study. All
of the patients had severe sciatica with or without low
back pain. The study was approved by the Institutional
Ethics Board, and written informed consent was autho-
rized by all patients. A total of 80 IVDs were collected
for bacterial culture, of which 10 were from levels L3–
L4, 41 were from L4–L5, and 29 were from L5–S1. The
patients received instrumented posterior lumbar fusion
after discectomy if they had spinal instability or spondy-
lolisthesis. The patients who had more severe lower back
pain than leg pain also underwent lumbar fusion None
of the patients had clinical or biological signs of infec-
tion before the operation. No patients were given antibi-
otics within one month prior to the surgery.
Preoperative imaging
For all patients, preoperative anteroposterior and lateral
lumbar spine radiographs were obtained in the standing
position. Magnetic resonance imaging (MRI) was
Page 2 of 5
performed for 68 patients. The Modic classification system
was applied to evaluate signal intensity changes in the adja-
cent vertebral body based on a previous protocol [8]. The
severity of disc degeneration was evaluated according to
the Pfirrmann system [9].
Intraoperative samples
During general anaesthesia induction, all patients re-
ceived prophylactic antibiotic treatment with a single
dose of cefazolin of 2 g following the instructions of the
Institutional Ethics Board. Strict aseptic techniques were
used for intraoperative disc removal. In brief, the skin of
the operation field was disinfected with povidone iodine
3 times, and then the antimicrobial incise drape (3 M
Ioban 2 Antimicrobial Incise Drape, 3 M Health Care,
St. Paul, MN, USA) was used to cover the surgical field.
Subsequently, the wound was irrigated twice using steril-
ized water before the discectomy of intervertebral discs.
Then, the specimen was handled only using a fresh in-
strument to minimize contamination and was quickly
transferred into a sterile pot and immediately covered
with the lid. Finally, tissue samples from the surrounding
muscles and ligaments were obtained at the same time
to serve as the contamination marker.
Microbiological culture
All of the tissue samples were sent to the Laboratory of
Bacteriology in our hospital. Under a class II laminar
flow safety cabinet, each sample was dissected into 5
segments with a sterilised scalpel. Three of the 5 seg-
ments were plated on three types of culture media
plates, and 2 of them were in enriched broth. For plate
culture, the dilacerated segments were first spread across
the surface of the plates, and then the tissue was embed-
ded into the plate. Briefly, the segments were embedded
in MacConkey agar plates that were aerobically incu-
bated at 37 °C (results read at 7 and 14 days), chocolate
agar plates incubated at 37 °C in 5% CO2 (results read at
7 and 14 days), and horse blood agar plates anaerobically
incubated at 37 °C (results read at 7 and 14 days),
respectively.
For broth enrichment, one segment was cultured in
cooked meat medium broth and incubated at 37 °C.
After 48 h of incubation, one broth subculture was
plated onto horse blood agar plates and anaerobically in-
cubated at 37 °C or onto chocolate agar plates and incu-
bated at 37 °C in 5% CO2, and the results were read 7
days later. A second segment was incubated in cooked
meat medium broth for 7 days and then plated onto
horse blood agar plates and anaerobically incubated at
37 °C or plated onto chocolate agar plates and incubated
at 37 °C in 5% CO2, and the results were read 7 days
later (14 days from surgery). When the samples had bac-
terial growth, they were identified with 16Sr DNA PCR
Tang et al. BMC Musculoskeletal Disorders (2018) 19:445 Page 3 of 5

with universe primers, and the products of PCR were Table 1 Relationships between bacteria and severity of disc
analysed and compared to known genes. degeneration
Disc degeneration total p
Statistical analysis Positive culture Negative culture
For continuous variables, the data were expressed as the Pfirrmann IV 3 15 18 0.162
mean ± SD. A two-sided Student’s t-test was used to ana- Pfirrmann V 20 39 59
lyse differences between the two groups. Categorical var-
Total 23* 54 77
iables were compared using Fisher’s exact test and the
*Three cases of suspicious contamination were excluded
Chi-square test. P < 0.05 was considered significantly
different.
pain. In this study, 32.5% of patients had low-virulence
Results bacteria in the IVDs, which was similar to previous re-
A total of 80 patients with 80 disc samples were enrolled ports of approximately 8%~ 53% [10]. In addition, the
in this study. The cohort included 35 males and 45 fe- surrounding muscles and ligaments served as contamin-
males, with a mean age of 51 ± 14.9 years. Among the pa- ation markers, so the isolated bacteria could be attrib-
tients, 17.6% had diabetes, and 36.8% were smokers. No uted to original growth inside the IVDs rather than
patients received preoperative transforaminal, epidural, or possible contamination. Therefore, we concluded that
facet joint injections before the surgery. No patients had a there was latent infection of low-virulence bacteria in
history of spinal or abdominal surgery. Of the patients, 12 IVDs with a prevalence of 28.75% after the exclusion of
received surgery at the level of L3/4, 40 underwent surgery three suspicious cases.
at L4/5, and 28 underwent surgery at L5/S1. In addition, the bacteria we isolated and identified
Microbiological disc cultures revealed that 26 disc were also similar to those in previous reports. In our
samples (26/80, 32.5%) were positive for bacterial study, P. acnes accounted for 26.25%, and CNS
growth. However, 3 cases had bacterial growth both in accounted for 6.25%. Stirling et al. first reported that the
discs and surrounding ligaments or muscles at the same prevalence of P. acnes and CNS was 44.4% (16/36) and
time, and the remaining 23 samples (23/80, 28.7%) only 5.5% (2/36) in degenerated IVDs, respectively [3]. Subse-
had bacterial growth in the discs. Considering that the 3 quently, several papers also reported the existence of
discs may have been contaminated during surgery, 23 low-virulence anaerobic bacteria in IVDs, especially P.
cases were considered absolute bacteria- positive. acnes, with the prevalence ranging from 45 to 84%.
As depicted in Table 1, P. acnes was detected in 21 sam- More importantly, the survival, reproduction and patho-
ples (21/80, 26.25%, one case had bacterium growth at both genicity of the bacterium was further validated in animal
the muscle and ligament), and coagulase-negative Staphylo- studies according to previous reports [11, 12]. Hence,
coccus (CNS) was detected in 5 samples (5/80, 6.25; two low-virulence anaerobic bacteria, especially P. acnes,
cases had bacteria growth at both the muscle and ligament). have been considered a new pathogenic factor for disc
Statistical analysis revealed that patients with bacteria- disease by increasing numbers of scholars.
positive in IVDs were significantly younger than
bacteria-negative patients (P = 0.03, Table 2). There were no Table 2 Patient and clinical characteristics in relation to
significant differences between positive and negative patients bacterial positivity
in smoking, diabetes and levels of discs (Table 2). Parameters n = 77* Positive culture Negative culture P
MRI revealed that 25 discs (25/80, 31.2%) had MCs n = 23* n = 54
(including Modic change I and Modic change II) at adja- Age 50.9 ± 12.1 44.7 ± 11.5 53.5 ± 11.5 0.003
cent vertebrae. Of these, 15 cases (15/80, 18.7%) were Male 35 13 22 0.203
bacteria-positive, while the remaining 10 cases (10/80,
Female 42 10 32
12.5%) were bacteria-negative. Table 3 shows that there
was a significant association between the presence of Diabetes 12 3 9 0.979
bacteria and MCs (P < 0.01). However, there was no stat- Smoker 26 6 20 0.352
istical association between bacterial infection and the se- VAS for back 4.2 ± 1.8 4.0 ± 1.7 4.2 ± 1.8 0.616
verity of disc degeneration (P = 0.162, Table 1). VAS for leg 5.1 ± 1.6 5.1 ± 1.5 5.1 ± 1.7 0.989
Level of disc culture
Discussion
L3/4 10 4 6 0.250
In recent years, multiple studies have reported
low-virulent anaerobic bacteria in disc material, suggest- L4/5 40 14 26
ing that such infection may contribute to degenerate L5/S1 27 5 22
spine conditions, including sciatica, neck pain, and back *Three cases of suspicious contamination were excluded
Tang et al. BMC Musculoskeletal Disorders (2018) 19:445
Table 3 Relationships between bacterial infection and MCs
Culture in discs total
Positive culture Negative culture
MCs (type I/ II) 15 10 25
No MCs 8 44 52
Total 23* 54 77
*Three cases of suspicious contamination were excluded
Despite this evidence, the hypothesis that latent infec-
tion of anaerobic low-virulence bacteria resided in IVDs
remains controversial. Several studies have suggested that
bacteria detected in disc cultures may originate from the
patient’s skin, the air, and laminar flow, especially that P.
acnes or CNS are normal flora residing on the human skin
[13, 14]. For example, McLorinan et al. demonstrated that
P. acnes would contaminate the incision during the oper-
ation, indicating that the bacteria was detected in 29.1% of
skin samples, 21.5% of tissue samples, and 16.5% of wash-
ings [14]. However, it is difficult to attribute all the isolated
bacteria to contamination because several papers have
demonstrated the existence of bacteria within the IVDs,
and the phylogroup patterns of bacteria were different
from those of skin [10].
Reports by Stirling and others suggest that low-
grade infection may play a role in inflammation and
herniation [3, 15–17]. In our study, positive cultures
were significantly associated with Modic changes in
the adjacent vertebrae, further supporting the theory
that MCs in the vertebrae adjacent to a previously
herniated disc may be related to oedema surrounding
an infected disc. Animal studies also demonstrated
that inoculation of anaerobic low-virulence bacteria
into IVDs would cause degenerative discogenic dis-
ease, such as Modic changes or disc degeneration,
which is different from the pyogenic discitis caused
by Staphylococcus aureus (S. aureus) [11, 12]. There-
fore, low-virulence anaerobic bacteria have been con-
sidered one of the reasons for Modic changes.
Unfortunately, there were still several limitations in this
study. First, all patients had to receive preoperative prophy-
lactic antibiotics according to the protocol of our Institu-
tional Ethics Board, which may have decreased the rate of
positive culture. Additionally, we acknowledge that the use
of antibiotics might also have reduced the number of posi-
tive cultures obtained from the ligament and muscle mater-
ial, which was interpreted in this study as a control for
contamination. Additionally, the polymerase chain reaction
was not performed directly on tissue to detect the bacterial
DNA. Although laboratory culture is the gold standard for
the diagnosis of bacterial pathogens, polymerase chain reac-
tion has advantages and disadvantages. Finally, more sam-
ples should have been included in the study to increase the
accuracy of the conclusions.
Page 4 of 5
Conclusion
p In summary, the results of our present study confirmed
previous findings of low-grade infection in patients
<0.001 undergoing surgery for a first time disc herniation. Such
infections might play important roles in the pathophysi-
ology of MCs and may change the therapeutic strategy
in the future.
Abbreviations
CNS: coagulase-negative Staphylococcus; IVDs: intervertebral discs;
MCs: Modic changes; P. acnes: Propionibacterium acnes
Acknowledgements
Not applicable.
Funding
This work was supported by grant from the National Natural Science Foundation
of China (NO. NSFC 81802180), which mainly contributed to the design of the
study. Also, this work was supported by grant from Natural Science Foundation of
Jiangsu Province (NO. BK20161274) and the Natural Science Foundation of
Suzhou (NO. kjxw2015056), both of which were responsible for sample collection,
bacterial culture, analysis of data and interpretation of results. Finally, this work
was supported by grant from the Science and Technology Bureau of Kunshan
(NO. KS1547), which played critical role in writing the manuscript.
Availability of data and materials
The datasets used and analysed during the current study are available from
the corresponding author upon reasonable request.
Authors’ contributions
The manuscript was approved by all authors. YC conceived anddesigned the
study and were responsible for the surgery; GQT acquired the data,
interpreted the results and involved in drafting and revising the manuscript;
ZW performed the radiological analysis and interpreted the results; JC
collected and transported the samples, acquired the epidemiological data of
patients and interpreted the results; ZSZ performed the bacterial culture and
identification; HBQ performed the statistical analysis and interpreted the
results during drafting and revising the manuscript.
Ethics approval and consent to participate
This study was approved by the Committee on Ethics and the Institutional
Review Board of Kunshan Hospital of Traditional Chinese Medicine, Nanjing
University of Chinese Medicine. Each patient signed a written informed
consent form before the study.
Consent for publication
Not applicable.
Competing interests
The authors declare that they have no competing interests.
Publisher’s Note
Springer Nature remains neutral with regard to jurisdictional claims in
published maps and institutional affiliations.
Received: 15 June 2018 Accepted: 6 December 2018
References
1. Modic MT, Ross JS. Lumbar degenerative disk disease. Radiology. 2007;245:
43–61.
2. Mulleman D, Mammou S, Griffoul I, Watier H, Goupille P. Pathophysiology of
disk-related sciatica. I.--evidence supporting a chemical component. Joint
Bone Spine. 2006;73:151–8.
3. Stirling A, Worthington T, Rafiq M, Lambert PA, Elliott TS. Association
between sciatica and Propionibacterium acnes. Lancet. 2001;357:2024–5.
4. Albert HB, Lambert P, Rollason J, Sorensen JS, Worthington T, Pedersen MB,
Norgaard HS, Vernallis A, Busch F, Manniche C, Elliott T. Does nuclear tissue
Tang et al. BMC Musculoskeletal Disorders (2018) 19:445 Page 5 of 5

infected with bacteria following disc herniations lead to Modic changes in

the adjacent vertebrae? Eur. Spine J. 2013;22:690–6.
5. Albert HB, Manniche C, Sorensen JS, Deleuran BW. Antibiotic treatment in
patients with low-back pain associated with Modic changes type 1 (bone
oedema): a pilot study. Br J Sports Med. 2008;42:969–73.
6. Jensen TS, Karppinen J, Sorensen JS, Niinimaki J, Leboeuf-Yde C. Vertebral
endplate signal changes (Modic change): a systematic literature review of
prevalence and association with non-specific low back pain. Eur Spine J.
2008;17:1407–22.
7. Urquhart DM, Zheng Y, Cheng AC, Rosenfeld JV, Chan P, Liew S, Hussain
SM, Cicuttini FM. Could low grade bacterial infection contribute to low back
pain? A systematic review. BMC Med. 2015;13:13.
8. Modic MT, Steinberg PM, Ross JS, Masaryk TJ, Carter JR. Degenerative disk
disease: assessment of changes in vertebral body marrow with MR imaging.
Radiology. 1988;166:193–9.
9. Pfirrmann CW, Metzdorf A, Zanetti M, Hodler J, Boos N. Magnetic resonance
classification of lumbar intervertebral disc degeneration. Spine Phila Pa.
2001;1976(26):1873–8.
10. Chen Z, Cao P, Zhou Z, Yuan Y, Jiao Y, Zheng Y. Overview: the role of
Propionibacterium acnes in nonpyogenic intervertebral discs. Int Orthop.
2016;40:1291–8.
11. Chen Z, Zheng Y, Yuan Y, Jiao Y, Xiao J, Zhou Z and Cao P. Modic changes
and disc degeneration caused by inoculation of Propionibacterium acnes
inside intervertebral discs of rabbits: a pilot study. Biomed Res Int 2016;
2016: 9612437.
12. Dudli S, Liebenberg E, Magnitsky S, Miller S, Demir-Deviren S, Lotz JC.
Propionibacterium acnes infected intervertebral discs cause vertebral bone
marrow lesions consistent with Modic changes. J Orthop Res. 2016;34:1447–55.
13. Ben-Galim P, Rand N, Giladi M, Schwartz D, Ashkenazi E, Millgram M, Dekel
S, Floman Y. Association between sciatica and microbial infection: true
infection or culture contamination? Spine (Phila Pa 1976). 2006;31:2507–9.
14. McLorinan GC, Glenn JV, McMullan MG, Patrick S. Propionibacterium acnes
wound contamination at the time of spinal surgery. Clin Orthop Relat Res.
2005:67–73.
15. Yuan Y, Chen Y, Zhou Z, Jiao Y, Li C, Zheng Y, Lin Y, Xiao J, Chen Z, Cao P.
Association between chronic inflammation and latent infection of
Propionibacterium acnes in non-pyogenic degenerated intervertebral discs:
a pilot study. Eur Spine J. 2017.
16. Dudli S, Miller S, Demir-Deviren S, Lotz JC. Inflammatory response of disc
cells against Propionibacterium acnes depends on the presence of lumbar
Modic changes. Eur Spine J. 2017.
17. Albert HB, Sorensen JS, Christensen BS and Manniche C. Antibiotic
treatment in patients with chronic low back pain and vertebral bone
edema (Modic type 1 changes): a double-blind randomized clinical
controlled trial of efficacy. Eur Spine J 2013; 22: 697–707.